Proper synaptic communication is critical for nervous system function and depends on neurotransmitter release through synaptic vesicle fusion to the presynaptic plasma membrane. This highly regulated process is mediated by a core membrane fusion machinery consisting of the SNARE proteins, and is further modulated by a number of accessory proteins. One such protein, complexin (Cpx), directly regulates SNARE-mediated synaptic vesicle fusion in a manner that is crucial for proper synaptic function. This proposal focuses on the mechanistic role that protein/lipid interactions play in complexin function. Cpx1 or Cpx2 single knockout mice display clear neurological and/or behavioral deficits, demonstrating the requirement of complexin's activities for proper neuronal function. Indeed, complete absence of Cpx function is lethal, as Cpx1/Cpx2 double knockout mice die shortly after birth. In worms and flies, Cpx knockout produces similarly severe neurological impairments. Cpx expression levels are altered in a variety of psychiatric and neurodegenerative diseases, including Huntington's, Parkinson's and Alzheimer's diseases, schizophrenia, depression, and bipolar disorder, and it has been suggested that altered Cpx expression levels may contribute to the symptomatology of these disorders. Cpx binds to the ternary SNARE complex and is known to directly regulate synaptic vesicle exocytosis. However, there has been great controversy regarding whether Cpx inhibits or stimulates synaptic vesicle fusion. The Cpx C-terminal domain (CTD) has been shown to mediate inhibition of spontaneous synaptic vesicle fusion in worms, and we have recently shown that binding of the CTD to phospholipid bilayers appears to be necessary for such inhibition. Our data suggest that synaptic vesicles are likely the relevant in vivo targets for the Cpx CTD, but it is unclear what might direct Cpx specifically to synaptic vesicles vs. other cellular membranes. We propose that membrane composition and curvature may play a critical role in targeting Cpx to vesicles. Additionally, the precise role of Cpx-vesicl interactions in Cpx function is unclear at present. We hypothesize that vesicle binding localizes Cpx to docking synaptic vesicles and thereby facilitates rapid interception of assembling SNARE complexes. To explore these hypotheses, I will study the interactions of the critical presynaptic protein complexin with phospholipid bilayers using powerful biophysical and biochemical techniques, and I will work to understand how this interaction contributes to synaptic function. Specifically, I will (1) determine how Cpx is directed to vesicles by characterizing the protein structural determinants of binding, and how lipid composition and curvature affect binding, and (2) assess how the energetics and kinetics of Cpx- lipid binding might influence Cpx function. These results will pave the way for critical subsequent in vivo studies to evaluate the resulting conclusions, facilitate increased understanding of Cpx function at the synapse, and contribute to an improved understanding of how alterations in complexin levels and/or function might affect synaptic function and synaptic biology in disease states.